Extracellular vesicles (EVs) are small lipid vesicles released by both prokaryotic and eukaryotic cells, involved in intercellular communication, immunomodulation and pathogenesis. In this study, we performed a characterization of the EVs produced by trophozoites of a clinical isolate of the free-living amoeba Naegleria fowleri (N. fowleri). Size distribution, zeta potential, protein profile and protease activity were analyzed. Under our incubation conditions, EVs of different sizes were observed, with a predominant population ranging from 206 to 227 nm. SDS-PAGE revealed protein bands of 25 to 260 KDa. The presence of antigenic proteins was confirmed by Western blot, which evidenced strongest recognition by rat polyclonal antibodies raised against N. fowleri in the region close to 80 KDa and included peptidases, as revealed by zymography. Proteins in selected immunorecognized bands were further identified using nano-ESI-MS/MS. A preliminary proteomic profile of the EVs identified at least 184 proteins as part of the vesicles’ cargo. Protease activity assays, in combination with the use of inhibitors, revealed the predominance of serine proteases. The present characterization uncovers the complexity of EVs produced by N. fowleri, suggesting their potential relevance in the release of virulence factors involved in pathogenicity. Owing to their cargo’s diversity, further research on EVs could reveal new therapeutic targets or biomarkers for developing rapid and accurate diagnostic tools for lethal infections such as the one caused by this amoeba.

Mancozeb is a worldwide fungicide used on a large scale in agriculture. The active component and its main metabolite, ethylene thiourea, has been related to health issues. Robust, fast, and reliable methodologies to quantify its presence in water are of great importance for environmental and health reasons. The electrochemical evaluation of mancozeb using a low-cost electrochemical electrode modified with poly (3,4-ethylene dioxythiophene), multi-walled carbon nanotubes, and gold nanoparticles is a novel strategy to provide an in-situ response for water pollution from agriculture. Additionally, the thermal-, electrochemical-, and photo-degradation of mancozeb and the production of ethylene thiourea under controlled conditions were evaluated in this research. The mancozeb solutions were characterized by electrochemical oxidation and ultraviolet-visible spectrophotometry, and the ethylene thiourea concentration was measured using ultra-high-performance liquid chromatography high-resolution mass spectrometry. The degradation study of mancozeb may provide routes for treatment in wastewater treatment plants. Therefore, a low-cost electrochemical electrode was fabricated to detect mancozeb in water with a robust electrochemical response in the linear range as well as a quick response at a reduced volume. Hence, our novel modified electrode provides a potential technique to be used in environmental monitoring for pesticide detection.

Negatively charged liposomes accomplished both functions as a reducing and stabilizing agent in the synthesis of gold nanotriangles (GNTs). Liposomes are based on a mixture of phospholipids phosphatidylcholine/phosphoglycerol, and they were used as a template phase to perform the GNTs. The method was evaluated under different conditions such as temperature, reaction time, phosphoglycerol chain length, and precursor concentration. Isotropic and anisotropic gold nanoparticles are formed simultaneously during the synthesis. Therefore, by combining centrifugation and depletion flocculation strategies, the sample was concentrated in terms of GNTs from 15% crude to 80% by using sodium dodecyl sulfate (SDS). As a result, a green colored dispersion was obtained containing highly purified, well-defined, negatively charged GNTs, where the edge length of most particles is centered in the range of 60–80 nm with an average thickness of 7.8 ± 0.1 nm. By this purification process, it was possible to highly increase the yield in terms of GNTs. Other surfactants [cetyltrimethylammonium chloride (CTAC), hexadecyltrimethylammonium bromide (CTAB), Tween 20, and dodecyldimethylammonium bromide] were evaluated during the purification stage, and both CTAB and CTAC show similar results to those obtained by using SDS. These GNTs are potential candidates for future applications in molecular imaging, photothermal therapy, drug delivery, biosensing, and photodynamic therapy.

Organic conductive polymers have been widely used as active layers in bioelectronic devices. In this work, a novel approach to entrap enzymes directly into the conductive active layer is described, using a polysaccharide as a surfactant. The surfactant allowed the electropolymerization from a micellar media and it acted as a doping agent in the conductive polymer. Gold nanotriangles were added to the matrix in order to enhance the enzymatic product quantification. The composition and oxidation state of the biocompatible conductive layer were confirmed by infrared spectrophotometric and Raman studies. Meanwhile, the gold nanotriangles presence, distribution and electrochemical activity were studied by transmission electron microscopy, atomic force microscopy, dynamic light scattering and cyclic voltammetry techniques. The inhibition of the enzyme, due to the presence of pesticides, was used to electrochemically quantify their concentration in real water samples. The concentration was confirmed by gas and liquid chromatography. Therefore, this novel composite active layer allows building a biosensor with suitable performance for an early warning in environmental control, especially in countries highly impacted by agricultural activities.

Graphite surfaces can be manipulated by several methods to create graphene structures of different shapes and sizes. Scanning tunneling microscopy (STM) can be used to create these structures either through mechanical contact between the tip and the surface or through electro-exfoliation. In the latter, the mechanisms involved in the process of exfoliation at an applied voltage are not fully understood. Here, we show how a graphite surface can be locally exfoliated in a systematic manner by applying an electrostatic force with a STM tip at the edge of a terrace, forming triangular flakes several nanometers in length. We demonstrate, through experiments and simulations, how these flakes are created by a two-step process: first a voltage ramp must be applied at the edge of the terrace, and then the tip must be scanned perpendicular to the edge. Ab initio electrostatic calculations reveal that the presence of charges on the graphite surface weakens the interaction between layers allowing for exfoliation at voltages in the same range as those used experimentally. Molecular dynamics simulations show that a force applied locally on the edge of a step produces triangular flakes such as those observed under STM. Our results provide new insights into surface modification that can be extended to other layered materials.